Recently, wild type ras genes have been shown to function as potent tumor suppressor genes. Both K-ras (hemizygous) and N-ras (homozygous) knockout mice are extremely prone to chemical induction of lung tumors and thymic lymphomas, respectively. Mice overexpressing wild type N-ras and rats overexpressing wild type K-ras and H-ras significantly inhibit thymic lymphomagenesis in mice and rat mammary tumorigenesis, respectively. These results indicate that wild type ras genes are tumor suppressors. This conclusion is further supported by observations of frequent loss of heterozygosity at the ras loci, which seem to fit the "two-hit" model of tumor suppressor gene inactivation in tumors. Another related tumor suppressor is RASSF1A that has been recently found to be the tumor suppressor residing on chromosome 3p21. RASSF1A is a Ras effector that binds to GTP-bound RAS. There are reports indicating that the occurrence of either K-ras mutations or RASSF1A inactivation activates the same signaling pathway in human tumors including lung. Accordingly, we hypothesize that activation of Ras-mediated pathways in mouse lung tumor progression occurs either by mutations of a Ras gene (oncogenic Ras and loss of wild type allele), or loss of function of the RASSF1A gene (LOH on Chr 3p21.3 and promoter hypermethylation), but infrequently by both, and the mechanisms used include the regulation of the ERK/AP-1 signaling pathways and activation of specific apoptosis inhibitors. Three specific aims are proposed to test our hypothesis: 1) to examine the mechanism of tumor suppressor function of wild type K-ras and RASSF1A using lung cancer cells conditionally expressing these genes and mouse embryonic fibroblasts lacking these genes; 2) to determine if K-ras alterations and RASSF1A hypermethylation occur independently in mouse lung tumorigenesis; and 3) to determine effect of RASSF1A deficiency and regulated expression of this gene on mouse lung tumor progression in mice. We propose that wild type K-ras and RASSF1A are related novel tumor suppressors of lung cancer as well as other cancer types. There is evidence to indicate that there are tissue-specific interactions between the wild type K-ras and RASSF1A in their roles as tumor suppressors. Understanding the mechanism(s) underlying these two newly discovered tumor suppressors would have immediate and important implications for our understanding of carcinogenesis and potentially provides better drug targets for cancer therapy.